Systemically applied nano-vaccines tend to accumulate in the liver. Within this organ, non-parenchymal cells (NPC), including Kupffer cells (KC), liver sinusoidal endothelial cells (LSEC), and dendritic cells (DC), induce T cell tolerance by default. Hence, unwanted uptake of nano-vaccines by tolerance-promoting liver NPC may limit the efficacy of DC-focused nano-vaccines for tumor therapy. However, appropriate adjuvants have previously been shown to reprogram KC and LSEC to induce T effector cells. Based on these findings, this project strives to improve nano-vaccination by identifying effective adjuvant combinations that activate antigen-presenting dendritic cells in secondary lymphoid organs and liver NPC to induce T effector cell responses. Since all of these cell populations express toll-like receptor 9, which is triggered by unmethylated CpG containing DNA oligodeoxynucleotides (ODN), we first conducted a comparative analysis of this type of adjuvants. In a second step, after identifying candidate CpG ODN, we co-applied siRNA targeting activation-induced anti-inflammatory proteins to enhance the immunogenic response. To this end, bone marrow-derived (BM)DC and liver NPC were differentially pretreated and examined by flow cytometry to assess the expression of surface activation markers. In addition, cytokine patterns of these cell populations were analyzed by Cytometric Bead Array (CBA). Furthermore, we conducted CD8+ T cell proliferation assays to assess the T cell stimulatory activity of DC and liver NPC. During these studies, CpG ODN SL03 was identified as the most potent stimulator for both liver NPC and BMDC. The addition of FOXO3 and STAT3-directed siRNA, respectively, further enhanced cytokine production by BMDC. Moreover, preliminary experiments indicate that accordingly stimulated BMDC in turn activate adjacent BMDC at a high extent. Ongoing experiments are dedicated to evaluating combinations of the aforementioned nucleic acid-based adjuvants when complexed with lipid-based nano-carriers and in combination with antigen-encoding mRNA.